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Human noroviruses (HuNoVs) are a diverse group of RNA viruses that cause endemic and pandemic acute viral gastroenteritis. Previously, we reported that many HuNoV strains require bile or bile acid (BA) to infect human jejunal intestinal enteroid cultures. BA was not essential for the replication of a pandemic-causing GII.4 HuNoV strain. We found the hydrophobic BA glycochenodeoxycholic acid (GCDCA) promotes the replication of the BA-dependent strain GII.3 in jejunal enteroids. Furthermore, we found that inhibition of the G-protein-coupled BA receptor, sphingosine-1-phosphate receptor 2 (S1PR2), by JTE-013, reduced GII.3 infection dose-dependently and inhibited GII.3 cellular uptake in enteroids. Herein, we sought to determine whether S1PR2 is required for other BA-dependent HuNoV strains, the BA-independent GII.4, and whether S1PR2 is required for BA-dependent HuNoV infection in HIEs from other small intestinal segments. We found a second S1PR2 inhibitor, GLPG2938, reduces GII.3 infection dose-dependently, and an S1PR2 agonist (CYM-5520) enhances GII.3 replication in the absence of GCDCA. GII.3 replication also is abrogated in the presence of JTE-013 and CYM-5520. JTE-013 inhibition of S1PR2 in jejunal HIEs reduces GI.1, GII.3, and GII.17 (BA-dependent) but not GII.4 Sydney (BA-independent) infection, providing additional evidence of strain-specific differences in HuNoV infection. Finally, GII.3 infection of duodenal, jejunal, and ileal lines derived from the same individual is reduced with S1PR2 inhibition, indicating a common mechanism of BA-dependent infection among multiple segments of the small intestine. Our results support a model where BA-dependent HuNoVs exploit BA effects on S1PR2 to infect the entire small intestine.IMPORTANCEHuman noroviruses (HuNoVs) are important viral human pathogens that cause both outbreaks and sporadic gastroenteritis. These viruses are diverse, and many strains are capable of infecting humans. Our previous studies have identified strain-specific requirements for hydrophobic bile acids (BAs) to infect intestinal epithelial cells. Moreover, we identified a BA receptor, sphingosine-1-phosphate receptor 2 (S1PR2), required for infection by a BA-dependent strain. To better understand how various HuNoV strains enter and infect the small intestine and the role of S1PR2 in HuNoV infection, we evaluated infection by additional HuNoV strains using an expanded repertoire of intestinal enteroid cell lines. We found that multiple BA-dependent strains, but not a BA-independent strain, all require S1PR2 for infection. In addition, BA-dependent infection requires S1PR2 in multiple segments of the small intestine. Together, these results indicate that S1PR2 has value as a potential therapeutic target for BA-dependent HuNoV infection.
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Ácidos e Sais Biliares , Norovirus , Receptores de Esfingosina-1-Fosfato , Replicação Viral , Humanos , Norovirus/efeitos dos fármacos , Norovirus/fisiologia , Norovirus/genética , Receptores de Esfingosina-1-Fosfato/metabolismo , Receptores de Esfingosina-1-Fosfato/antagonistas & inibidores , Replicação Viral/efeitos dos fármacos , Ácidos e Sais Biliares/farmacologia , Ácidos e Sais Biliares/metabolismo , Infecções por Caliciviridae/virologia , Infecções por Caliciviridae/metabolismo , Piridinas/farmacologia , Gastroenterite/virologia , Jejuno/virologia , Jejuno/metabolismo , Organoides/virologia , Organoides/metabolismo , PirazóisRESUMO
In vitro models, such as primary cells and continuous cell lines routinely used for evaluating drug candidates, have limitations in their translational relevance to human diseases. Organotypic cultures are increasingly being used to assess therapeutics for various cancers and infectious diseases. Monitoring drug cytotoxicity in cell cultures is crucial in drug development, and several commercially available kits for cytotoxicity assessment offer distinct advantages and limitations. Given the complexity of organoid cultures, including donor-driven variability, we investigated drug-treated, tissue stem cell-derived human intestinal organoid responses with commonly used cell cytotoxicity assay kits. Using seven different compounds, we compared the cytotoxicity assay performance of two different leaky membrane-based and two metabolism-based assays. Significant variability was seen in reported viability outcomes across assays and organoid lines. High baseline activity of lactate dehydrogenase (LDH) in four human intestinal organoid lines required modification of the standard LDH assay protocol. Additionally, the LDH assay reported unique resilience to damage in a genetically-modified line contrasting results compared to other assays. This study highlights factors that can impact the measurement of cell cytotoxicity in intestinal organoid models, which are emerging as valuable new tools for research and pre-clinical drug testing and suggest the need for using multiple assay types to ensure reliable cytotoxicity assessment.
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L-Lactato Desidrogenase , Organoides , Humanos , Organoides/efeitos dos fármacos , Organoides/metabolismo , Organoides/citologia , L-Lactato Desidrogenase/metabolismo , Sobrevivência Celular/efeitos dos fármacos , Intestinos/citologia , Mucosa Intestinal/efeitos dos fármacos , Mucosa Intestinal/citologia , Mucosa Intestinal/metabolismoRESUMO
Human noroviruses (HuNoVs) are the leading cause of acute gastroenteritis worldwide. Currently, there are no targeted antivirals for the treatment of HuNoV infection. Histo-blood group antigens (HBGAs) on the intestinal epithelium are cellular attachment factors for HuNoVs; molecules that block the binding of HuNoVs to HBGAs thus have the potential to be developed as antivirals. Human milk oligosaccharides (HMOs) are glycans in human milk with structures analogous to HBGAs. HMOs have been shown to act as decoy receptors to prevent the attachment of multiple enteric pathogens to host cells. Previous X-ray crystallography studies have demonstrated the binding of HMO 2'-fucosyllactose (2'FL) in the same pocket as HBGAs for some HuNoV strains. We evaluated the effect of 2'FL on the replication of a globally dominant GII.4 Sydney [P16] HuNoV strain using human intestinal enteroids (HIEs) from adults and children. A significant reduction in GII.4 Sydney [P16] replication was seen in duodenal and jejunal HIEs from multiple adult donors, all segments of the small intestine from an adult organ donor and in two pediatric duodenal HIEs. However, 2'FL did not inhibit HuNoV replication in two infant jejunal HIEs that had significantly lower expression of α1-2-fucosylated glycans. 2'FL can be synthesized in large scale, and safety and tolerance have been assessed previously. Our data suggest that 2'FL has the potential to be developed as a therapeutic for HuNoV gastroenteritis.
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BACKGROUND: The in vitro cultivation of human noroviruses allows a comparison of antibody levels measured in neutralization and histoblood group antigen (HBGA)-blocking assays. METHODS: Serum samples collected during the evaluation of an investigational norovirus vaccine (HIL-214 [formerly TAK-214]) were assayed for neutralizing antibody levels against the vaccine's prototype Norwalk virus/GI.1 (P1) virus strain. Results were compared to those previously determined using HBGA-blocking assays. RESULTS: Neutralizing antibody seroresponses were observed in 83% of 24 vaccinated adults, and antibody levels were highly correlated (r=0.81, P<0.001) with those measured by HBGA-blocking. CONCLUSIONS: GI.1-specific HBGA-blocking antibodies are a surrogate for neutralization of GI.1 norovirus.
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Human noroviruses (HuNoVs) are a significant cause of both epidemic and sporadic acute gastroenteritis worldwide. The lack of a reproducible culture system for HuNoVs was a major obstacle in studying virus replication and pathogenesis for almost a half-century. This barrier was overcome with our successful cultivation of multiple HuNoV strains in human intestinal enteroids (HIEs), which has significantly advanced HuNoV research. We previously optimized culture media conditions and generated genetically-modified HIE cultures to enhance HuNoV replication in HIEs. Building upon these achievements, we now present additional advancements to this culture system, which involve testing different media, unique HIE lines, and additional virus strains. HuNoV infectivity was evaluated and compared in new HIE models, including HIEs generated from different intestinal segments of individual adult organ donors, HIEs made from human embryonic stem cell-derived human intestinal organoids that were transplanted into mice (H9tHIEs), genetically-engineered (J4 FUT2 knock-in [ KI ], J2 STAT1 knock-out [ KO ]) HIEs, as well as HIEs derived from a patient with common variable immunodeficiency (CVID) and from infants. Our findings reveal that small intestinal HIEs, but not colonoids, from adults, H9tHIEs, HIEs from a CVID patient, and HIEs from infants support HuNoV replication with segment and strain-specific differences in viral infection. J4 FUT2-KI HIEs exhibit the highest susceptibility to HuNoV infection, allowing the cultivation of a broader range of GI and GII HuNoV strains than previously reported. Overall, these results contribute to a deeper understanding of HuNoVs and highlight the transformative potential of HIE cultures in HuNoV research. Importance: Human noroviruses (HuNoVs) are very contagious and cause significant acute gastroenteritis globally, but studying them has been hindered by the lack of a reproducible culture system for nearly 50 years. This barrier was overcome by successfully cultivating multiple HuNoV strains in human intestinal enteroids (HIEs), advancing HuNoV research. We previously optimized culture conditions and developed genetically modified HIEs to enhance HuNoV replication. In this study, we tested different media, unique HIE lines, and additional virus strains, evaluating HuNoV infectivity in new HIE models. These models include HIEs from various intestinal segments of adult donors, human embryonic stem cell-derived HIEs transplanted into mice (H9tHIEs), genetically-engineered HIEs (J4 FUT2 knock-in [ KI ], J2 STAT1 knock-out [ KO ]), HIEs from a common variable immunodeficiency (CVID) patient, and from infants. Our findings show that adult small intestinal HIEs, H9tHIEs, CVID patient HIEs, and infant HIEs support HuNoV replication with segment and strain-specific differences. J4 FUT2-KI HIEs exhibited the highest susceptibility, allowing cultivation of a broader range of HuNoV strains. These results enhance the understanding of HuNoVs and highlight the transformative potential of HIE cultures in HuNoV research.
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Human noroviruses, globally the main cause of viral gastroenteritis, show strain specific affinity for histo-blood group antigens (HBGA) and can successfully be propagated ex vivo in human intestinal enteroids (HIEs). HIEs established from jejunal stem cells of individuals with different ABO, Lewis and secretor geno- and phenotypes, show varying susceptibility to such infections. Using bottom-up glycoproteomic approaches we have defined and compared the N-linked glycans of glycoproteins of seven jejunal HIEs. Membrane proteins were extracted, trypsin digested, and glycopeptides enriched by hydrophilic interaction liquid chromatography and analyzed by nanoLC-MS/MS. The Byonic software was used for glycopeptide identification followed by hands-on verifications and interpretations. Glycan structures and attachment sites were identified from MS2 spectra obtained by higher-energy collision dissociation through analysis of diagnostic saccharide oxonium ions (B-ions), stepwise glycosidic fragmentation of the glycans (Y-ions), and peptide sequence ions (b- and y-ions). Altogether 694 unique glycopeptides from 93 glycoproteins were identified. The N-glycans encompassed pauci- and oligomannose, hybrid- and complex-type structures. Notably, polyfucosylated HBGA-containing glycopeptides of the four glycoproteins tetraspanin-8, carcinoembryonic antigen-related cell adhesion molecule 5, sucrose-isomaltase and aminopeptidase N were especially prominent and were characterized in detail and related to donor ABO, Lewis and secretor types of each HIE. Virtually no sialylated N-glycans were identified for these glycoproteins suggesting that terminal sialylation was infrequent compared to fucosylation and HBGA biosynthesis. This approach gives unique site-specific information on the structural complexity of N-linked glycans of glycoproteins of human HIEs and provides a platform for future studies on the role of host glycoproteins in gastrointestinal infectious diseases.
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Antígenos de Grupos Sanguíneos , Infecções por Caliciviridae , Fucose , Glicoproteínas , Antígenos de Histocompatibilidade , Jejuno , Organoides , Glicômica , Proteômica , Genótipo , Fenótipo , Glicoproteínas/química , Glicoproteínas/genética , Glicoproteínas/metabolismo , Fucose/metabolismo , Glicosilação , Antígenos de Grupos Sanguíneos/química , Antígenos de Grupos Sanguíneos/genética , Antígenos de Grupos Sanguíneos/metabolismo , Antígenos de Histocompatibilidade/química , Antígenos de Histocompatibilidade/genética , Antígenos de Histocompatibilidade/metabolismo , Humanos , Glicopeptídeos/química , Infecções por Caliciviridae/sangue , Infecções por Caliciviridae/imunologia , Infecções por Caliciviridae/metabolismo , Organoides/metabolismo , Jejuno/metabolismo , Jejuno/virologiaRESUMO
INTRODUCTION: A surge of human influenza A(H7N9) cases began in 2016 in China from an antigenically distinct lineage. Data are needed about the safety and immunogenicity of 2013 and 2017 A(H7N9) inactivated influenza vaccines (IIVs) and the effects of AS03 adjuvant, prime-boost interval, and priming effects of 2013 and 2017 A(H7N9) IIVs. METHODS: Healthy adults (n = 180), ages 19-50 years, were enrolled into this partially blinded, randomized, multicenter phase 2 clinical trial. Participants were randomly assigned to 1 of 6 vaccination groups evaluating homologous versus heterologous prime-boost strategies with 2 different boost intervals (21 vs 120 days) and 2 dosages (3.75 or 15â µg of hemagglutinin) administered with or without AS03 adjuvant. Reactogenicity, safety, and immunogenicity measured by hemagglutination inhibition and neutralizing antibody titers were assessed. RESULTS: Two doses of A(H7N9) IIV were well tolerated, and no safety issues were identified. Although most participants had injection site and systemic reactogenicity, these symptoms were mostly mild to moderate in severity; injection site reactogenicity was greater in vaccination groups receiving adjuvant. Immune responses were greater after an adjuvanted second dose, and with a longer interval between prime and boost. The highest hemagglutination inhibition geometric mean titer (95% confidence interval) observed against the 2017 A(H7N9) strain was 133.4 (83.6-212.6) among participants who received homologous, adjuvanted 3.75â µg + AS03/2017 doses with delayed boost interval. CONCLUSIONS: Administering AS03 adjuvant with the second H7N9 IIV dose and extending the boost interval to 4 months resulted in higher peak antibody responses. These observations can broadly inform strategic approaches for pandemic preparedness. Clinical Trials Registration. NCT03589807.
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Anticorpos Antivirais , Imunização Secundária , Subtipo H7N9 do Vírus da Influenza A , Vacinas contra Influenza , Influenza Humana , Vacinas de Produtos Inativados , Humanos , Vacinas contra Influenza/imunologia , Vacinas contra Influenza/administração & dosagem , Vacinas contra Influenza/efeitos adversos , Adulto , Masculino , Feminino , Pessoa de Meia-Idade , Subtipo H7N9 do Vírus da Influenza A/imunologia , Vacinas de Produtos Inativados/imunologia , Vacinas de Produtos Inativados/administração & dosagem , Vacinas de Produtos Inativados/efeitos adversos , Anticorpos Antivirais/sangue , Influenza Humana/prevenção & controle , Influenza Humana/imunologia , Adulto Jovem , Esquemas de Imunização , Testes de Inibição da Hemaglutinação , Estados Unidos , Imunogenicidade da Vacina , Anticorpos Neutralizantes/sangue , Polissorbatos/administração & dosagem , Polissorbatos/efeitos adversos , alfa-Tocoferol/administração & dosagem , alfa-Tocoferol/efeitos adversos , Esqualeno/administração & dosagem , Esqualeno/efeitos adversos , Esqualeno/imunologia , Voluntários Saudáveis , Combinação de Medicamentos , Adjuvantes de Vacinas/administração & dosagem , Vacinação/métodos , Adjuvantes Imunológicos/administração & dosagem , Adjuvantes Imunológicos/efeitos adversosRESUMO
Human noroviruses (HuNoVs) are a diverse group of RNA viruses that cause both endemic and pandemic acute viral gastroenteritis. Previously we reported that many strains of HuNoV require bile or bile acid (BA) to infect human jejunal intestinal enteroid cultures. Of note, BA was not essential for replication of a pandemic-causing GII.4 HuNoV strain. Using the BA-requiring strain GII.3, we found that the hydrophobic BA GCDCA induces multiple cellular responses that promote replication in jejunal enteroids. Further, we found that chemical inhibition of the G-protein coupled receptor, sphingosine-1- phosphate receptor 2 (S1PR2), by JTE-013 reduced both GII.3 infection in a dose- dependent manner and cellular uptake in enteroids. Herein, we sought to determine if S1PR2 is required by other BA-dependent HuNoV strains and BA-independent GII.4, and if S1PR2 is required for BA-dependent HuNoV infection in other segments of the small intestine. We found JTE-013 inhibition of S1PR2 in jejunal HIEs reduces GI.1, GII.3, and GII.17 (BA-dependent) but not the GII.4 Sydney variant (BA-independent) infection, providing additional evidence of strain-specific differences in HuNoV infection. GII.3 infection of duodenal, jejunal and ileal lines derived from the same individual was also reduced with S1PR2 inhibition, indicating a common mechanism of BA-dependent infection among multiple segments of the small intestine. Our results support a model where BA-dependent HuNoV exploit the activation of S1PR2 by BA to infect the entire small intestine. Importance: Human noroviruses (HuNoVs) are important viral human pathogens that cause both outbreaks and sporadic gastroenteritis. These viruses are diverse, and many strains are capable of infecting humans. Our previous studies have identified strain-specific requirements for hydrophobic bile acids (BAs) to infect intestinal epithelial cells. Moreover, we identified a BA receptor, sphingosine-1-phosphate receptor 2 (S1PR2), required for infection by a BA-dependent strain. To better understand how various HuNoV strains enter and infect the small intestine and the role of S1PR2 in HuNoV infection, we evaluated infection by additional HuNoV strains using an expanded repertoire of intestinal enteroid cell lines. We found that multiple BA-dependent strains, but not a BA- independent strain, all required S1PR2 for infection. Additionally, BA-dependent infection required S1PR2 in multiple segments of the small intestine. Together these results indicate S1PR2 has value as a potential therapeutic target for BA-dependent HuNoV infection.
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BACKGROUND: Risk factors for ciprofloxacin or MDR in primary care urine specimens are not well defined. OBJECTIVES: We created a primary care-specific antibiogram for Escherichia coli isolates from cases with complicated and uncomplicated urinary tract infection (UTI) and evaluated risk factors for ciprofloxacin, trimethoprim/sulfamethoxazole and MDR among Enterobacterales. METHODS: We conducted a cross-sectional study to determine resistance and risk factors by collecting urine cultures from all patients (≥18 years) presenting with provider-suspected UTI at two primary care, safety-net clinics in Houston, TX, USA between November 2018 and March 2020. RESULTS: Among 1262 cultures, 308 cultures grew 339 uropathogens. Patients with Enterobacterales (nâ=â199) were mostly female (93.5%) with a mean age of 48.5 years. E. coli was the predominant uropathogen isolated (nâ=â187/339; 55%) and had elevated trimethoprim/sulfamethoxazole (43.6%) and ciprofloxacin (29.5%) resistance, low nitrofurantoin (1.8%) resistance, and no fosfomycin resistance. Among E. coli, 10.6% were ESBL positive and 24.9% had MDR. Birth outside the U.S.A., prior (2 year) trimethoprim/sulfamethoxazole resistance, and diabetes mellitus were associated with trimethoprim/sulfamethoxazole resistance. Prior (60 day) fluoroquinolone use, prior ciprofloxacin resistance and both diabetes mellitus and hypertension were strongly associated with ciprofloxacin resistance. Prior fluoroquinolone use and a history of resistance to any studied antibiotic were associated with MDR, while pregnancy was protective. CONCLUSIONS: We found elevated resistance to UTI-relevant antimicrobials and novel factors associated with resistance; these data can be incorporated into clinical decision tools to improve organism and drug concordance.
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Diabetes Mellitus , Gammaproteobacteria , Gravidez , Humanos , Feminino , Pessoa de Meia-Idade , Masculino , Ciprofloxacina/farmacologia , Estudos Transversais , Escherichia coli , Combinação Trimetoprima e Sulfametoxazol/farmacologia , Fatores de Risco , Fluoroquinolonas , Testes de Sensibilidade Microbiana , Resistência a Múltiplos Medicamentos , Atenção Primária à SaúdeRESUMO
Background: Studies have demonstrated low hepatitis A virus (HAV) vaccination rates among persons with HIV (PWH). Methods: We conducted a retrospective study of persons entering HIV care at two clinics in Houston, Texas between 2010 and 2018. We defined those eligible for HAV vaccination as those who had no history of HAV vaccination and had a negative anti-HAV IgG at entry to care. Kaplan-Meier curves summarized time to receipt of HAV vaccines. The proportions of patients who received 1 and 2 HAV vaccines at 6, 12, and 24 months were estimated. Cox proportional hazards regression evaluated associations between patient characteristics and vaccination. Significant factors were included in a multivariable Cox proportional hazards model. Results: Of 6,515 patients, 1372 were eligible for HAV vaccination. Of eligible patients, 29.2 % received 1 HAV vaccination at 6 months, 37.1 % at 12 months, and 47.8 % at 24 months. At 6 months, 10 % received 2 HAV vaccinations, 21.1 % at 12 months, and 33.4 % at 24 months. In multivariable analysis, men who have sex with men (adjusted HR 1.35, 95 % CI 1.06, 1.73) or those who had CD4 count ≥ 200 cells/µl (adjusted HR 2.52, 95 % CI 1.89, 3.37) had their second vaccination sooner than those who were not men who have sex with men or who had CD4 counts < 200 cells/µl, respectively. Patients > 50 years of age had their second vaccination sooner than those aged 30-50 years (adjusted HR 1.47, 95 % CI 1.08, 1.99). Those with active substance history had a longer time to second vaccination compared to those with no substance use history (adjusted HR 0.57, 95 % CI 0.40, 0.82). Conclusions: HAV vaccination rates were low and highlight the need for effective solutions to address HAV immunization gaps in PWH, especially among young patients, those with active substance use disorders, and those with significant immunocompromise.
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Background: Studies have demonstrated low hepatitis B virus (HBV) vaccine series completion among persons with human immunodeficiency virus (HIV). Methods: We conducted a retrospective record review of persons entering HIV care at 2 clinics in Houston, Texas, between 2010 and 2018. Kaplan-Meier curves summarized time to receipt of HBV vaccines for those eligible for vaccination. We estimated the proportions of patients who had received 1, 2, or 3 HBV vaccine doses at 12 and 24 months after entry to care. A Prentice Williams and Peterson total time model was used to evaluate associations between patient characteristics and time to vaccination. Results: Of the 5357 patients who entered care, 2718 were eligible for HBV vaccination. After 2 years of follow-up, 51.2% of those eligible had received 1 HBV vaccine, 43.2% had received 2, and 28.4% received 3 vaccines. With adjustment for significant cofactors, patients whose CD4 cell count was ≥200/µL (adjusted hazard ratio [aHR], 1.43 [95% confidence interval (CI), 1.29-1.59]) and transgender patients (1.49 [1.08-2.04]) received any given vaccine dose sooner than those with CD4 cell counts <200/µL or cisgender patients, respectively. Compared with non-Hispanic whites, Hispanic patients were vaccinated sooner (aHR, 1.28 [95% CI, 1.07-1.53]). Those with an active substance use history had a significantly longer time to vaccination than those with no substance use history (aHR, 0.73 [95% CI, .62-.85]). Conclusions: Strategies are needed to increase HBV vaccine completion rates in our study population, particularly among those with CD4 cell counts <200/µL or with a substance use disorder.
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A GII.2 outbreak in an efficacy study of a bivalent virus-like particle (VLP) norovirus vaccine, TAK-214, in healthy US adults provided an opportunity to examine GII.4 homotypic vs. GII.2 heterotypic responses to vaccination and infection. Three serological assays (VLP-binding, histoblood group antigen-blocking, and neutralizing) were performed for each genotype. Results were highly correlated within a genotype but not between genotypes. Although the vaccine provided protection from GII.2-associated disease, little GII.2-specific neutralization occurred after vaccination. Choice of antibody assay can affect assessments of human norovirus vaccine immunogenicity.
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Human noroviruses (HuNoVs) are the leading cause of acute gastroenteritis. In immunocompetent hosts, symptoms usually resolve within 3 days; however, in immunocompromised persons, HuNoV infection can become persistent, debilitating, and sometimes life-threatening. There are no licensed therapeutics for HuNoV due to a near half-century delay in its cultivation. Treatment for chronic HuNoV infection in immunosuppressed patients anecdotally includes nitazoxanide, a broad-spectrum antimicrobial licensed for treatment of parasite-induced gastroenteritis. Despite its off-label use for chronic HuNoV infection, nitazoxanide has not been clearly demonstrated to be an effective treatment. In this study, we standardized a pipeline for antiviral testing using multiple human small intestinal enteroid lines representing different intestinal segments and evaluated whether nitazoxanide inhibits replication of five HuNoV strains in vitro. Nitazoxanide did not exhibit high selective antiviral activity against any HuNoV strain tested, indicating it is not an effective antiviral for HuNoV infection. Human intestinal enteroids are further demonstrated as a model to serve as a preclinical platform to test antivirals against HuNoVs to treat gastrointestinal disease. Abstr.
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Infecções por Caliciviridae , Gastroenterite , Norovirus , Humanos , Gastroenterite/tratamento farmacológico , Antivirais/farmacologia , Antivirais/uso terapêutico , Padrões de Referência , Infecções por Caliciviridae/tratamento farmacológico , Replicação ViralRESUMO
Acute gastroenteritis caused by human noroviruses (HuNoVs) is a significant global health and economic burden and is without licensed vaccines or antiviral drugs. The GII.4 HuNoV causes most epidemics worldwide. This virus undergoes epochal evolution with periodic emergence of variants with new antigenic profiles and altered specificity for histo-blood group antigens (HBGA), the determinants of cell attachment and susceptibility, hampering the development of immunotherapeutics. Here, we show that a llama-derived nanobody M4 neutralizes multiple GII.4 variants with high potency in human intestinal enteroids. The crystal structure of M4 complexed with the protruding domain of the GII.4 capsid protein VP1 revealed a conserved epitope, away from the HBGA binding site, fully accessible only when VP1 transitions to a "raised" conformation in the capsid. Together with dynamic light scattering and electron microscopy of the GII.4 VLPs, our studies suggest a mechanism in which M4 accesses the epitope by altering the conformational dynamics of the capsid and triggering its disassembly to neutralize GII.4 infection.
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Antígenos de Grupos Sanguíneos , Infecções por Caliciviridae , Norovirus , Humanos , Proteínas do Capsídeo/química , Capsídeo/metabolismo , Norovirus/genética , Sítios de Ligação , Epitopos/metabolismo , Antígenos de Grupos Sanguíneos/metabolismoRESUMO
Vaccine protection against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection wanes over time, requiring updated boosters. In a phase 2, open-label, randomized clinical trial with sequentially enrolled stages at 22 US sites, we assessed safety and immunogenicity of a second boost with monovalent or bivalent variant vaccines from mRNA and protein-based platforms targeting wild-type, Beta, Delta and Omicron BA.1 spike antigens. The primary outcome was pseudovirus neutralization titers at 50% inhibitory dilution (ID50 titers) with 95% confidence intervals against different SARS-CoV-2 strains. The secondary outcome assessed safety by solicited local and systemic adverse events (AEs), unsolicited AEs, serious AEs and AEs of special interest. Boosting with prototype/wild-type vaccines produced numerically lower ID50 titers than any variant-containing vaccine against all variants. Conversely, boosting with a variant vaccine excluding prototype was not associated with decreased neutralization against D614G. Omicron BA.1 or Beta monovalent vaccines were nearly equivalent to Omicron BA.1 + prototype or Beta + prototype bivalent vaccines for neutralization of Beta, Omicron BA.1 and Omicron BA.4/5, although they were lower for contemporaneous Omicron subvariants. Safety was similar across arms and stages and comparable to previous reports. Our study shows that updated vaccines targeting Beta or Omicron BA.1 provide broadly crossprotective neutralizing antibody responses against diverse SARS-CoV-2 variants without sacrificing immunity to the ancestral strain. ClinicalTrials.gov registration: NCT05289037 .
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Vacinas contra COVID-19 , COVID-19 , Humanos , Vacinas contra COVID-19/efeitos adversos , SARS-CoV-2/genética , COVID-19/prevenção & controle , Anticorpos Amplamente NeutralizantesRESUMO
We compared the serologic responses of 1 dose versus 2 doses of a variant vaccine (Moderna mRNA-1273 Beta/Omicron BA.1 bivalent vaccine) in adults. A 2-dose boosting regimen with a variant vaccine did not increase the magnitude or the durability of the serological responses compared to a single variant vaccine boost.
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Vacina de mRNA-1273 contra 2019-nCoV , Adulto , Humanos , Vacinas Combinadas , Protocolos Clínicos , RNA Mensageiro/genéticaRESUMO
As part of a multicenter study evaluating homologous and heterologous COVID-19 booster vaccines, we assessed the magnitude, breadth, and short-term durability of binding and pseudovirus-neutralizing antibody (PsVNA) responses following a single booster dose of NVX-CoV2373 in adults primed with either Ad26.COV2.S, mRNA-1273, or BNT162b2 vaccines. NVX-CoV2373 as a heterologous booster was immunogenic and associated with no safety concerns through Day 91. Fold-rises in PsVNA titers from baseline (Day 1) to Day 29 were highest for prototypic D614G variant and lowest for more recent Omicron sub-lineages BQ.1.1 and XBB.1. Peak humoral responses against all SARS-CoV-2 variants were lower in those primed with Ad26.COV2.S than with mRNA vaccines. Prior SARS CoV-2 infection was associated with substantially higher baseline PsVNA titers, which remained elevated relative to previously uninfected participants through Day 91. These data support the use of heterologous protein-based booster vaccines as an acceptable alternative to mRNA or adenoviral-based COVID-19 booster vaccines. This trial was conducted under ClinicalTrials.gov: NCT04889209.
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In this brief report, we compare the magnitude and durability of the serologic response of one versus two doses (separated by 56 days) of a variant vaccine (Moderna mRNA-1273 Beta/Omicron BA.1 bivalent vaccine) in adults.
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Human noroviruses (HuNoVs) are the leading cause of acute gastroenteritis. In immunocompetent hosts, symptoms usually resolve within three days; however, in immunocompromised persons, HuNoV infection can become persistent, debilitating, and sometimes life-threatening. There are no licensed therapeutics for HuNoV due to a near half-century delay in its cultivation. Treatment for chronic HuNoV infection in immunosuppressed patients anecdotally includes nitazoxanide, a broad-spectrum antimicrobial licensed for treatment of parasite-induced gastroenteritis. Despite its off-label use for chronic HuNoV infection, nitazoxanide has not been clearly demonstrated to be an effective treatment. In this study, we established a standardized pipeline for antiviral testing using multiple human small intestinal enteroid (HIE) lines representing different intestinal segments and evaluated whether nitazoxanide inhibits replication of 5 HuNoV strains in vitro . Nitazoxanide did not exhibit high selective antiviral activity against any HuNoV strains tested, indicating it is not an effective antiviral for norovirus infection. HIEs are further demonstrated as a model to serve as a pre-clinical platform to test antivirals against human noroviruses to treat gastrointestinal disease.